Isolation of a Bacillus safensis from mine tailings in Peru, genomic characterization and characterization of its cyanide- degrading enzyme CynD

Cyanide is widely used in industry as a potent lixiviant due to its capacity to tightly bind metals. This property imparts cyanide enormous toxicity to all known organisms. Thus, industries that utilize this compound must reduce its concentration in recycled or waste waters. Physical, chemical, and biological treatments have been used for cyanide remediation; however, none of them meet all the desired characteristics: efficiency, low cost and low environmental impact. A better understanding of metabolic pathways and biochemistry of enzymes involved in cyanide degradation is a necessary step to improve cyanide bioremediation efficacy to satisfy the industry requirements. Here, we used several approaches to explore this topic. We have isolated three cyanide-degrading Bacillus strains from water in contact with mine tailings from Lima, Peru, and classified them as Bacillus safensis PER-URP-08, Bacillus licheniformis PER-URP-12, and Bacillus subtilis PER-URP-17 based on 16S rRNA gene sequencing and core genome analyses. Additionally, core genome analyses of 132 publicly available genomes of Bacillus pumilus group including B. safensis and B. altitudinis allowed us to reclassify some strains and identify two strains that did not match with any known species of the Bacillus pumilus group. We searched for possible routes of cyanide-degradation in the genomes of these three strains and identified putative B. licheniformis PER-URP-12 and B. subtilis PER-URP-17 rhodaneses and B. safensis PER-URP-08 cyanide dihydratase (CynD) sequences possibly involved cyanide degradation. We identified characteristic C-terminal residues that differentiate CynD from B. pumilus and B. safensis, and showed that, differently from CynD from B. pumilus C1, recombinant CynD from the Bacillus safensis PER-URP-08 strain remains active up to pH 9 and presents a distinct oligomerization pattern at pH 8 and 9. Moreover, transcripts of B. safensis PER-URP-08 CynD (CynDPER-URP-08) are strongly induced in the presence of cyanide. Our results warrant further investigation of B. safensis PER-URP-08 and CynDPER-URP-08 as potential tools for cyanide-bioremediation.

Cyanides and Their Attenuation in Wastewaters Using Green Chemistry

Cyanides, though naturally occurring, are environmental pollutants when not treated properly. Some methods used to attenuate cyanides in waste waters from industrial processes are based simply on changing the physico-chemical properties of the waste water such as the pH and temperature. The effectiveness of these methods are based on hydrolysis of the cyanide and volatilization of the hydrogen cyanide formed. Another reaction which takes place simultaneously is ultraviolet-catalysed oxidation which converts the cyanide to bicarbonates and carbonates.The changes in the cyanide degradation rate approaches a minimum faster if the cyanide solution is maintained at a higher than ambient constant temperature.

Effect of cultural conditions on the growth and linamarase production by a local species of Lactobacillus fermentum isolated from cassava effluent

Background This study was designed to investigate the effect of cultural conditions on growth and production of linamarase by a local species of Lactobacillus fermentum isolated from cassava effluent. Isolation and identification of bacteria from cassava effluent were carried out using the culture-dependent method and polyphasic taxonomy, respectively, while screening for cyanide degradation, and the effects of cultural conditions on the growth and linamarase activity of L. fermentum were investigated based on standard procedures. Results A total of twenty-one bacterial isolates were obtained from cassava effluent, and isolate MA 9 had the highest growth of 2.8 × 1010 cfu/ml in minimum medium, confirmed as safe, identified as Lactobacillus fermentum and selected for further study. The highest growth of 2.498 OD and linamarase activity of 2.49 U/ml were observed at inoculums volume of 0.10 ml at 48-h incubation period, while optimum growth of 1.926 OD and linamarase activity of 1.66 U/ml occurred at pH 5.5. At 37 °C, the optimum growth of 0.34 OD was recorded with the highest linamarase activity of 0.81 U/ml at 30 °C. However, the incubation period of 48 h stimulated an optimum growth of 3.091 OD with corresponding linamarase activity of 1.81 U/ml, while the substrate concentration of 400 ppm favours a maximum growth of 2.783 OD with linamarase activity of 1.86 U/ml at 48 h of incubation. The supplemented of 10 mM calcium ions stimulated optimum linamarase activity of 2.65 U/ml. Conclusion Lactobacillus fermentum could be used as starter culture in cassava fermentation for the production cassava-based product with reduced cyanide content.

Enhancement of PMS activation in an UV/ozone process for cyanide degradation: a comprehensive study

Purpose This paper aims to study peroxymonosulfate (PMS) activation in the ultraviolet (UV)/ozone process for toxic cyanide degradation from aqueous solution by a novel and simple method. Design/methodology/approach Photocatalytic degradation of cyanide (CN-) was carried out using a bench-scale photoreactor. Optimization of the UV/ozone process for the highest removal of cyanide was obtained. The effect of parameters such as ozone concentration, PMS concentration, temperature, cations (Cu2+, Co2+ and Fe2+), cyanide concentration, anions (bicarbonate, carbonate, chloride, nitrite, nitrate and sulfate [SO42−]) and scavengers (ethanol [EtOH], humic acid, TBA and NaN3) was investigated for CN- degradation. Findings Complete removal of 50 mg/L cyanide was obtained in 4 min in an ozone/UV/PMS process. The cyanide removal increased from 49.3% to 100% by adding the persulfate dosage up to 100 mg/L. The effect of various cations (II) on the cyanide degradation was enhanced in the order Cu2+ > Co2+ > Fe2+. Hydroxyl radical based on different radical quenchers such as salicylic acid proved as the main oxidizing radical for oxidation. The application of ozone/UV/PMS to treat wastewater containing cyanide shows high degradation efficiency. Research limitations/implications The ozone/UV/PMS system could be a process for degradation and detoxification of cyanide. Practical implications This study provided a simple and effective method for degradation of cyanide from aqueous solution. This method was applicable to protect environment from a huge amount of toxic cyanide wastewater produced by different industrial processes. Originality/value The PMS activation is done via a simple and effective method, which is carried out with the ozone/UV system. There are two main innovations. One is that the novel catalytic role of bimetallic ions in the ozone reaction with cyanide and the further decomposition of intermediate products is investigated. The other is that the optimized conditions were obtained for the removal of cyanide as a water contaminant. Furthermore, predominant oxidizing species by PMS activation are identified.

Ethylene Biosynthesis Inhibition Combined with Cyanide Degradation Confer Resistance to Quinclorac in Echinochloa crus-galli var. mitis

Echinochloa crus-galli var. mitis has rarely been reported for herbicide resistance, and no case of quinclorac resistance has been reported so far. Synthetic auxin-type herbicide quinclorac is used extensively to control rice weeds worldwide. A long history of using quinclorac in Chinese rice fields escalated the resistance in E. crus-galli var. mitis against this herbicide. Bioassays in Petri plates and pots exhibited four biotypes that evolved into resistance to quinclorac ranking as JS01-R > AH01-R > JS02-R > JX01-R from three provinces of China. Ethylene production in these biotypes was negatively correlated with resistance level and positively correlated with growth inhibition. Determination of the related ethylene response pathway exhibited resistance in biotypes that recorded a decline in 1-aminocyclopropane-1-carboxylic acid (ACC) content, ACC synthase oxidase activities, and less inducible ACS and ACO genes expressions than the susceptible biotype, suggesting that there was a positive correlation between quinclorac resistance and ethylene biosynthesis inhibition. Cyanides produced during the ethylene biosynthesis pathway mainly degraded by the activity of β-cyanoalanine synthase (β-CAS). Resistant biotypes exhibited higher β-CAS activity than the susceptible ones. Nucleotide changes were found in the EcCAS gene of resistant biotypes as compared to sensitive ones that caused three amino acid substitutions (Asn-105-Lys, Gln-195-Glu, and Gly-298-Val), resulting in alteration of enzyme structure, increased binding residues in the active site with its cofactor, and decreased binding free energy; hence, its activity was higher in resistant biotypes. Moreover, these mutations increased the structural stability of the enzyme. In view of the positive correlation between ethylene biosynthesis inhibition and cyanide degradation with resistance level, it is concluded that the alteration in ethylene response pathway or at least variation in ACC synthase and ACC oxidase enzyme activities—due to less relative expression of ACS and ACO genes and enhanced β-CAS activity, as well as mutation and increased relative expression of EcCAS gene—can be considered as a probable mechanism of quinclorac resistance in E. crus-galli var. mitis.